Quinolinone derivative and pharmaceutical composition for preventing or treating allergic diseases such as asthma or atopic dermatitis including the quinolinone derivative as active ingredient

ABSTRACT

The present invention relates to a novel quinolinone derivative compound that regulates intracellular signal transduction mediated by TSLP and IL-33 to exhibit efficacy in preventing or treating allergic diseases such as asthma or atopic dermatitis. The quinolinone derivative compound of the present invention can effectively suppress inflammatory responses of allergic diseases such as asthma or atopic dermatitis. The present invention also relates to a pharmaceutical composition including the quinolinone derivative compound. The pharmaceutical composition of the present invention can be used to fundamentally prevent or treat various allergic and asthmatic diseases.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a U.S. National Stage Application of InternationalApplication No. PCT/KR2018/000165, filed on Jan. 4, 2018, which claimsthe benefit under 35 USC 119(a) and 365(b) of Korean Patent ApplicationNo. 10-2017-0002326, filed on Jan. 6, 2017 and Korean Patent ApplicationNo. 10-2018-0001019, filed on Jan. 4, 2018, in the Korean IntellectualProperty Office, the entire disclosure of which is incorporated hereinby reference for all purposes.

TECHNICAL FIELD

The present invention relates to a novel quinolinone derivative, andmore specifically to a 4- or 5-substituted2-aryl-oxazolo[4,5-c]quinolin-4(5H)-one as a quinolinone derivativecompound that regulates signal transduction mediated by TSLP and IL-33to exhibit efficacy in preventing or treating allergic diseases such asasthma or atopic dermatitis, and a pharmaceutical composition forpreventing or treating allergic diseases such as asthma or atopicdermatitis including the quinolinone derivative compound as an activeingredient.

BACKGROUND ART

Treatment of allergic inflammatory diseases with bronchodilators andanti-inflammatory agents is based on allopathy. These therapeutic agentsare temporarily effective in ameliorating symptoms of allergic diseasesbut cannot basically control allergic diseases, failing to fundamentallytreat the diseases.

Environmental diseases such as bronchial asthma, atopic skin diseases,and allergic rhinitis are known as immune diseases and Th2 cells arewell known to play a pivotal role in causing allergic responses. Whenstimulated by an antigen in lymphocytes, CD4 T cells can differentiateinto various types of Th cells depending on cytokines recognizedsimultaneously by the cells. When the recognized cytokines are type 2cytokines, such as thymic stromal lymphopoietin (TSLP) or IL-4, suchcells differentiate into Th2 to cause allergic responses.

Interleukin-33 (IL-33) is an innate cytokine produced mainly by mucosalepithelial cells when various external stimuli are applied. IL-33 isknown to play an important role in regulating allergic immune responses,mainly asthma, mediated by Th2 cells. The IL-33 receptor complex forIL-33-mediated signaling consists of IL-33 as a ligand, ST2 (IL-1R4) asa ligand binder, and IL-1 receptor accessory protein (IL-IRAcP) as asignal transducer. Th2 inflammatory cytokines, including IL-4, IL-5,IL-6, IL-13 and IL-8, and chemokines are produced by stimulation ofIL-33. When bound with IL-33, the IL-33 receptor complex activatesmolecules of downstream signaling systems such as NF-kB and AP-1 throughIL-1 receptor-associated kinase (IRAK), TNF receptor associated factor 6(TRAF6), and/or MAPKs.

Overall, it is expected that allergic diseases can be basically treatedby the regulation of TSLP and IL-33, cytokines that play important rolesin the differentiation of Th2 cells.

PRIOR ART DOCUMENTS

-   Non-Patent Document 1: Julia V, Macia L, Dombrowicz D. Nat Rev    Immunol. 2015; 15:308-322.-   Non-Patent Document 2: S. Kamijo, H. Takeda, T. Tokura, M.    Suzuki, K. Inui, M. Hara, H. Matsuda, A. Matsuda, K. Oboki, T.    Ohno, H. Saito, S. Nakae, K. Sudo, H. Suto, S. Ichikawa, H.    Ogawa, K. Okumura, T. Takai, IL-33-mediated innate response and    adaptive immune cells contribute to maximum responses of protease    allergen-induced allergic airway inflammation, J Immunol, 190 (2013)    4489-4499.

DETAILED DESCRIPTION OF THE INVENTION Problems to be Solved by theInvention

Thus, the present invention intends to provide a novel quinolinonederivative that regulates intracellular signal transduction mediated byTSLP and IL-33 to exhibit efficacy in preventing or treating allergicdiseases such as asthma or atopic dermatitis.

The present invention also intends to provide a pharmaceuticalcomposition for preventing or treating allergic diseases such as asthmaor atopic dermatitis including the novel quinolinone derivative as anactive ingredient and a pharmaceutically acceptable carrier.

Means for Solving the Problems

One aspect of the present invention provides a quinolinone derivativerepresented by Formula 1a or 1b:

A description will be given concerning the structures and substituentsof Formulae 1a and 1b and specific examples of the quinolinonederivatives.

The present invention also provides a pharmaceutical composition forpreventing or treating allergic diseases such as asthma or atopicdermatitis including the novel derivative represented by Formula 1a or1b as an active ingredient and a pharmaceutically acceptable carrier.

Effects of the Invention

The quinolinone derivative compound of the present invention caneffectively suppress inflammatory responses of allergic diseases such asasthma or atopic dermatitis.

In addition, the pharmaceutical composition of the present invention canbe used to fundamentally prevent or treat various allergic diseases suchas asthma or atopic dermatitis.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a histogram showing the inhibitory effect of the compound ofFormula 2 according to the present invention on TSLP-TSLPR interaction,which was evaluated by ELISA.

FIG. 2 is a histogram showing the inhibitory effect of the compound ofFormula 7 according to the present invention on TSLP-TSLPR interaction,which was evaluated by ELISA.

FIG. 3 is a histogram showing the inhibitory effect of the compound ofFormula 10 according to the present invention on IL-6 secretion by IL-33inhibition.

FIGS. 4 and 5 show the numbers of total cells and eosinophils in theairways of house dust mite (HDM)-induced allergic airway inflammationmouse models treated with the compounds of Formula 5 (KB-1517) andFormula 10 (KB-1518) (200 μg each) according to the present invention toinvestigate the effects of the compounds.

FIG. 6 shows the chemical shift perturbations of Thr 120 of IL-33protein by compounds of the present invention.

FIG. 7 shows the chemical shift perturbations of Glu 121 of IL-33protein by compounds of the present invention.

FIG. 8 shows the treatment schedule with the Formula 5 (KB 1517) andFormula 10 (KB 1518) (200 μg each) according to the present invention inhouse dust mite (HDM)-induced allergic airway inflammation mouse models.

BEST MODE FOR CARRYING OUT THE INVENTION

The present invention will now be described in more detail.

The inventors of the present invention have found that quinolinonederivatives represented by Formulae 1a and 1b effectively inhibit TSLPand IL-33, key cytokines inducing allergic diseases such as asthma oratopic dermatitis, from binding to their receptors. The presentinvention has been accomplished based on this finding.

One aspect of the present invention is directed to a quinolinonederivative represented by Formula 1a or 1b:

wherein R₁ and R₂ may be identical to or different from each other andare each independently selected from C₁-C₄ alkyl groups, C₁-C₄ alkylenegroups, and —Z-alkyl (wherein Z is a heteroatom selected from O, S, andN or is —(CH₂)_(m)— wherein m is an integer from 0 to 5), X and Y areeach independently selected from hydrogen (H), halogen groups (fluoro(F), chloro (Cl), bromo (Br), and iodo (I)), a hydroxyl group (OH), anamino group (NH₂), a nitro group (NO₂), a cyano group (CN), atrifluoromethyl group (CF₃), CO—R′ (wherein R′ is a C₁-C₄ alkyl group),NHR′ (wherein R′ is a C₁-C₄ alkyl group), NR′R′ (wherein each R′ isindependently a C₁-C₄ alkyl group), NHCOR′ (wherein R′ is a C₁-C₄ alkylgroup), and COOR′ (wherein R′ is a C₁-C₄ alkyl group), p is an integerfrom 0 to 4, provided that when p is an integer of 2 or more, the Ygroups may be identical to or different from each other, and q is aninteger from 0 to 5, provided that when q is an integer of 2 or more,the X groups may be identical to or different from each other,

wherein R₁, R₂, X, Y, p, and q are as defined in Formula 1a.

The quinolinone derivative represented by Formula 1a or 1b regulatesintracellular signal transduction mediated by TSLP and IL-33 andinhibits the binding of thymic stromal lymphopoietin (TSLP) to TSLPreceptor or the binding of IL-33 to ST-2.

Specifically, the quinolinone derivative represented by Formula 1a or 1bmay be selected from, but not limited to, the compounds of Formulae 2 to15:

The quinolinone derivative of the present invention can be preparedthrough a series of successive reactions: Sandmeyer reaction,Suzuki-coupling reaction in the presence of a palladium catalyst, Heckreaction, nitro reduction, cyclization under basic conditions, andsubstitution. For example, the quinolinone derivative represented byFormula 1a or 1b may be prepared by the following procedure. First,ethyl 2-chlorooxazole-4-carboxylate is obtained by Sandmeyer reaction ofethyl 2-aminooxazole-4-carboxylate as a starting material. Then, theethyl 2-chlorooxazole-4-carboxylate is allowed to react with asubstituted arylboronic acid to synthesize a 2-aryloxazole-4-carboxylatederivative. Subsequently, the 2-aryloxazole-4-carboxylate derivative issubjected to Heck reaction with 2-iodo-1-nitrobenzene to prepare asubstituted 2-aryl-5-aryloxazole-4-carboxylate derivative. The nitrogroup of the substituted 2-aryl-5-aryloxazole-4-carboxylate derivativeis reduced by hydrogenation. An intramolecular reaction between theester group and the amine group under basic conditions and a subsequentsubstitution reaction with dialkylethyl bromide under basic conditionsgive the quinolinone derivative represented by Formula 1a or 1b.

More specifically, the compounds of Formulae 2 to 13 can be synthesizedby the following reaction schemes:

Reaction scheme [1] schematically shows a synthetic route to thequinolinone derivative of Formula 1a or 1b wherein both R₁ and R₂ aremethyl groups, and Reaction schemes [2] and [3] schematically showsynthetic routes to the quinolinone derivative of Formula 1a or 1bwherein both R₁ and R₂ are ethyl groups.

A further aspect of the present invention is directed to apharmaceutical composition for preventing or treating an allergicdisease such as asthma or atopic dermatitis including the quinolinonederivative represented by Formula 1a or 1b as an active ingredient. Thepharmaceutical composition of the present invention regulatesintracellular signal transduction mediated by TSLP and IL-33 andinhibits the binding of thymic stromal lymphopoietin (TSLP) to TSLPreceptor or the binding of IL-33 to ST-2.

The pharmaceutical composition of the present invention can be used toprevent and treat a wide range of allergic diseases such as asthma oratopic dermatitis. Examples of the allergic diseases include, but notlimited to, atopic dermatitis, urticarial rhinitis, allergic rhinitis,and asthmatic diseases.

The pharmaceutical composition of the present invention may be complexedwith other known drugs for the prevention and treatment of allergicdiseases such as asthma or atopic dermatitis before administration ormay further include one or more other additives selected from carriers,diluents, adjuvants, and stabilizers.

The dosage form of the composition according to the present inventionmay vary depending on the mode of administration desired. Examples ofsuch dosage forms include, but not limited to, solid, semi-solid, andliquid formulations, such as tablets, pills, powders, capsules, gels,ointments, emulsions, and suspensions. The composition of the presentinvention may be administered in unit dosage forms suitable for singleadministration of precise dosages. The composition of the presentinvention may be administered orally or parenterally. For parenteraladministration, the composition of the present invention may beadministered intravenously, subcutaneously or intramuscularly.

Depending on the formulation desired, the composition may furtherinclude one or more pharmaceutically acceptable carriers, diluents,adjuvants, and stabilizers, which are defined as aqueous-based vehiclescommonly used to formulate pharmaceutical compositions for humanadministration.

The term “carrier” means a substance that facilitates the incorporationof a compound into cells or tissues. Examples of suitable carriersinclude, but not limited to, carbohydrate-based compounds, such aslactose, amylose, dextrose, sucrose, sorbitol, mannitol, starch, andcellulose, gum acacia, calcium phosphate, alginate, gelatin, calciumsilicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose,water, syrups, salt solutions, alcohols, gum Arabic, vegetable oils,such as corn oil, cotton seed oil, soybean oil, olive oil, and coconutoil, polyethylene glycol, methyl cellulose, methyl hydroxybenzoate,propyl hydroxybenzoate, talc, magnesium stearate, and mineral oils,which are commonly used to formulate pharmaceutical compositions. Theterm “diluent” is defined as a substance diluted in water that candissolve the compound of interest as well as stabilize the biologicallyactive form of the compound. Examples of suitable diluents includedistilled water, physiological saline, Ringer's solution, dextrosesolution, and Hank's solution. The stabilizers can be selected from thegroup consisting of proteins, carbohydrates, buffers, and mixturesthereof. The composition of the present invention may optionally furtherinclude one or more additives. Examples of such optional additivesinclude, but not limited to, lubricating agents, wetting agents,sweetening agents, flavoring agents, emulsifying agents, suspendingagents, and preservatives.

Such additional additives as carriers, diluents, adjuvants, andstabilizers may be used in amounts effective to acquire pharmaceuticallyacceptable formulations in view of the solubility, biological activity,and other characteristics of the active ingredient.

As used herein, the term “prevent” or “preventing” refers to inhibitinga disease or disorder from occurring in an animal or human that may bepredisposed to the disease or disorder but has not yet been diagnosed ashaving it. As used herein, the term “treat” or “treating” refers toinhibiting the development of a disease or disorder or ameliorating oreliminating the disease or disorder.

As used herein, the term “including as an active ingredient” means thepresence of the corresponding ingredient in an amount necessary orsufficient to achieve a desired biological effect. In real applications,the active ingredient is used in a therapeutically effective amount totreat a target disease and such an amount can suitably be determinedtaking into consideration other toxicities caused by the activeingredient. For example, the amount of the active ingredient may varydepending on various factors, such as the disease or condition to betreated, the dosage form of the composition, the size of a subject orthe severity of the disease or condition. The effective amount of thecomposition can be empirically determined by those skilled in the artwithout excessive experiments.

By “pharmaceutically acceptable” is meant that the biological activityand properties of the compound are not impaired.

Other terms and abbreviations used herein may be understood as theirmeanings recognized generally by those skilled in the art, unlessotherwise defined.

MODE FOR CARRYING OUT THE INVENTION

The present invention will be explained in more detail with reference tothe following examples. These examples are provided to assist inunderstanding the invention and are not intended to limit the scope ofthe invention.

Synthesis Examples: Synthesis of Inventive Quinolinone Derivatives

The inventive quinolinone derivatives of Formulae 2-15 were synthesizedaccording to reaction schemes [1], [2], and [3].

(a) Ethyl 2-chlorooxazole-4-carboxylate (21) as intermediate

Ethyl 2-aminooxazole-4-carboxylate (468 mg, 3 mmol) was added tot-butylnitrite (540 μl, 0.45 mmol) and copper chloride (600 mg, 4.5mmol) in acetonitrile (22 ml) at 60° C. The mixture was heated at 80° C.for 1 h. After cooling, the mixture was partitioned withdichloromethane, ice, and hydrochloric acid. The aqueous layer wasfurther extracted with dichloromethane. The organic layer was washedwith brine, dried over MgSO₄, and evaporated. The crude product waspurified by silica gel column chromatography (hexane/Et₂O, 7:1 to 4:1,v/v) to afford Intermediate 21 (338 mg, 64%) as a fluffy white solid.

R_(f)=0.38 (hexane/Et₂O=2:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.20(s, 1H), 4.40 (q, J=7.2 Hz, 2H), 1.39 (t, J=6.9 Hz, 3H). LRMS (ESI) m/z176.1 [M+H]⁺.

(b) Synthesis of Ethyl 2-Aryloxazole-4-Carboxylate Derivatives asIntermediates Through Suzuki Reaction

(1) Ethyl 2-chlorooxazole-4-carboxylate (257 mg, 1.47 mmol),4-fluorophenylboronic acid (252 mg, 1.8 mmol, 1.2 eq.), andtetrakis(triphenylphosphine)palladium(0) (85 mg, 0.07 mmol, 0.05 eq.)were dissolved in toluene (20 mL) and a 2 M potassium carbonate solution(2.0 mL, 4.0 mmol) under a nitrogen atmosphere at room temperature. Thesolution was refluxed with stirring for 1 h. After cooling to roomtemperature, the reaction mixture was partitioned with ethyl acetate anda 2 M sodium hydroxide solution. The aqueous layer was washed twice withethyl acetate. The organic layer washed with brine, dried over MgSO₄,and concentrated in vacuo. The crude product was purified by silica gelcolumn chromatography (hexane/Et₂O, 5:1 to 3:1, v/v) to affordIntermediate 22b (250 mg, 73%) as a fluffy white solid.

R_(f)=0.30 (hexane/Et₂O=2:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.25(s, 1H), 8.11 (dd, J=5.4 and 8.9 Hz, 2H), 7.16 (t, J=9.0 Hz, 2H), 4.42(q, J=7.2 Hz, 2H), 1.40 (t, J=7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ ppm166.2, 162.9, 161.7, 161.3, 143.7, 134.7, 129.2, 129.1, 122.8, 122.8,116.3, 116.1, 116.0, 115.7, 61.4, 14.3. LRMS (ESI) m/z 257.8 [M+Na]⁺.HRMS (ESI) m/z calculated for C₁₂H₁₀FNO₃Na⁺ [M+Na]⁺: 258.0537; found:258.0528.

(2) Ethyl 2-phenyloxazole-4-carboxylate (22a)

R_(f)=0.30 (hexane/Et₂O=2:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.28(s, 1H), 8.12 (dd, J=2.1 and 7.2 Hz, 2H), 7.16 (dd, J=1.8 and 5.3 Hz,2H), 4.43 (q, J=7.2 Hz, 2H), 1.41 (t, J=7.2 Hz, 3H). LRMS (ESI) m/z218.0 [M+H]⁺ and 239.9 [M+Na]⁺.

(3) Ethyl 2-(4-chlorophenyl)oxazole-4-carboxylate (22c)

R_(f)=0.38 (hexane/Et₂O=2:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.27(s, 1H), 8.05 (d, J=8.4 Hz, 2H), 7.45 (d, J=7.2 Hz, 2H), 4.43 (q, J=7.2Hz, 2H), 1.40 (t, J=7.2 Hz, 3H). LRMS (ESI) m/z 251.8 [M+H]⁺ and 273.8[M+Na]⁺.

(4) Ethyl 2-(4-(trifluoromethyl)phenyl)oxazole-4-carboxylate (22d)

R_(f)=0.38 (hexane/Et₂O=2:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.32(s, 1H), 8.24 (d, J=8.1 Hz, 2H), 7.74 (d, J=8.1 Hz, 2H), 4.45 (q, J=7.2Hz, 2H), 1.42 (t, J=7.2 Hz, 3H). LRMS (ESI) m/z 286.0 [M+H]⁺ and 308.1[M+Na]⁺.

(5) Ethyl 2-(4-methoxyphenyl)oxazole-4-carboxylate (22e)

R_(f)=0.13 (hexane/Et₂O=2:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.24(s, 1H), 8.06 (d, J=9.0 Hz, 2H), 6.98 (d, J=9.0 Hz, 2H), 4.43 (q, J=7.2Hz, 2H), 1.42 (t, J=7.2 Hz, 3H). LRMS (ESI) m/z 247.7 [M+H]⁺ and 269.9[M+Na]⁺.

(c) Synthesis of ethyl 2-aryl-5-aryloxazole-4-carboxylate Derivatives asIntermediates by Heck Reaction

(1) A mixture of 22b (192 mg, 0.8 mmol), 2-iodonitrobenzene (398 mg, 1.6mmol, 2.0 eq.), palladium acetate (11.2 mg, 0.05 mmol, 0.06 eq.),triphenylphosphine (21 mg, 0.08 mmol, 0.1 eq.), cesium carbonate (651.6mg, 2.0 mmol, 2.5 eq.), and DMF (4 mL) was flushed with nitrogen andheated at 140° C. for 3 h. After cooling, the reaction mixture wasdiluted with ethyl acetate, washed with water and brine, dried overMgSO₄, and concentrated in vacuo. The crude product was purified bysilica gel column chromatography (hexane/Et₂O, 5:1 to 1:1, v/v) toafford Intermediate 23b (192 mg, 67%) as a yellow needle-like crystal.

R_(f)=0.35 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.20(d, J=8.1 Hz, 1H), 8.14 (q, J=8.4 Hz, 1H), 8.18-8.09 (m, 1H), 7.83-7.65(m, 3H), 7.19 (t, J=8.7 Hz, 2H), 4.34 (q, J=7.2 Hz, 2H), 1.29 (t, J=7.2Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 166.3, 161.3, 160.6, 151.0,148.5, 132.9, 132.6, 131.3, 130.2, 129.3, 129.2, 124.9, 122.6, 122.5,122.5, 116.3, 116.0, 61.6, 14.0. LRMS (ESI) m/z 357.4 [M+H]⁺, 379.0[M+Na]⁺, and 395.0 [M+K]⁺. HRMS (ESI) m/z calculated for C₁₈H₁₄FN₂O₅ ⁺[M+H]⁺: 357.0881; found: 357.0865.

(2) Ethyl 5-(2-nitrophenyl)-2-phenyloxazole-4-carboxylate (23a)

R_(f)=0.23 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.19(d, J=7.8 Hz, 1H), 8.16-8.08 (m, 1H), 8.12 (d, J=7.8 Hz, 1H), 7.76-7.66(m, 1H), 7.77 (d, J=4.2 Hz, 2H), 7.55-7.43 (m, 3H), 4.32 (q, J=7.2 Hz,2H), 1.27 (t, J=7.2 Hz, 3H). LRMS (ESI) m/z 339.3 [M+H]⁺, 361.1 [M+Na]⁺,and 377.0 [M+Na]⁺. HRMS (ESI) m/z calculated for C₁₈H₁₄N₂O₅Na⁺ [M+Na]⁺:361.0795; found: 361.0778.

(3) Ethyl 2-(4-chlorophenyl)-5-(2-nitrophenyl)oxazole-4-carboxylate(23c)

R_(f)=0.30 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.18(d, J=8.1 Hz, 1H), 8.05 (dd, J=4.2 and 10.1 Hz, 1H), 8.05 (d, J=8.7 Hz,1H), 7.81-7.65 (m, 3H), 7.46 (d, J=8.7 Hz, 1H), 7.46 (dd, J=4.2 and 9.3Hz, 1H), 4.31 (q, J=7.2 Hz, 2H), 1.26 (t, J=7.2 Hz, 3H); ¹³C NMR (75MHz, CDCl₃) δ ppm 161.2, 160.5, 151.1, 148.5, 137.6, 132.8, 132.6,131.3, 130.3, 129.2, 128.6, 124.9, 124.6, 122.6, 61.6, 29.7, 14.0. LRMS(ESI) m/z 373.1 [M+H]⁺, 395.0 [M+Na]⁺, and 411.0 [M+k]⁺. HRMS (ESI) m/zcalculated for C₁₈H₁₄ClN₂O₅ ⁺ [M+H]⁺: 373.0586; found: 373.0589.

(4) Ethyl5-(2-nitrophenyl)-2-(4-(trifluoromethyl)phenyl)oxazole-4-carboxylate(23d)

R_(f)=0.20 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.24(d, J=8.1 Hz, 2H), 8.21 (d, J=10.5 Hz, 1H), 7.86-7.68 (m, 5H), 4.32 (q,J=7.2 Hz, 2H), 1.27 (t, J=7.2 Hz, 3H); ¹³C NMR (75 MHz, CDCl₃) δ ppm166.1.1, 159.9, 151.7, 148.5, 133.0, 132.6, 127.3, 126.0, 125.9, 124.9,61.8, 14.0. LRMS (ESI) m/z 407.0 [M+H]⁺, 428.7 [M+Na]⁺, and 445.3[M+K]⁺. HRMS (ESI) m/z calculated for C₁₈H₁₄F₃N₂O₅ ⁺ [M+H]⁺: 407.0849;found: 407.0809.

(5) Ethyl 2-(4-methoxyphenyl)-5-(2-nitrophenyl)oxazole-4-carboxylate(23e)

R_(f)=0.20 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.18(d, J=8.7 Hz, 1H), 8.07 (d, J=8.7 Hz, 2H), 7.82-7.64 (m, 3H), 7.00 (t,J=9.0 Hz, 2H), 4.33 (q, J=7.2 Hz, 2H), 1.28 (t, J=7.2 Hz, 3H); ¹³C NMR(75 MHz, CDCl₃) δ ppm 162.1, 161.5, 161.5, 150.4, 148.5, 132.8, 132.6,131.1, 130.0, 128.8, 124.8, 122.8, 118.8, 114.3, 61.5, 55.5, 29.7, 14.1.LRMS (ESI) m/z 369.1 [M+H]⁺, 391.1 [M+Na]⁺, and 407.1 [M+K]⁺. HRMS (ESI)m/z calculated for C₁₉H₁₇N₂O₆ ⁺ [M+H]⁺: 369.1081; found: 369.1090.

(d) Synthesis of Amine Derivatives as Intermediates ThroughHydrogenation (1) Ethyl5-(2-aminophenyl)-2-(4-fluorophenyl)oxazole-4-carboxylate (24b)

A palladium catalyst was added to a solution of 23b (192 mg, 0.54 mmol)in MeOH (15 mL). The mixture was stirred under a hydrogen atmosphere (50psi) for 1 h. The reaction mixture was filtered through a bed of Celiteand volatiles were removed by reduction to afford Intermediate 24b (145mg, 82%) as a yellow solid.

R_(f)=0.30 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm8.18-8.07 (m, 1H), 8.12 (q, J=8.4 Hz, 1H), 7.43 (dd, J=1.2 and 7.8 Hz,1H), 7.29 (td, J=8.1 and 1.5 Hz, 1H), 7.22-7.08 (m, 1H), 7.16 (t, J=8.4Hz, 1H), 6.84 (td, J=7.8 and 1.2 Hz, 1H), 6.82 (d, J=8.1 Hz, 1H), 4.39(q, J=7.2 Hz, 2H), 4.16 (bs, 2H), 1.34 (t, J=7.2 Hz, 3H); ¹³C NMR (75MHz, CDCl₃) δ ppm 166.2, 162.8, 162.1, 159.8, 154.4, 145.7, 131.8,131.6, 129.8, 129.1, 129.0, 122.8, 122.8, 118.2, 116.7, 116.3, 116.0,112.7, 61.5, 14.2. LRMS (ESI) m/z 327.1 [M+H]⁺ and 349.1 [M+Na]⁺. HRMS(ESI) m/z calculated for C₁₈H₁₆FN₂O₃ ⁺ [M+H]⁺: 327.1139; found:327.1133.

(2) Ethyl 5-(2-aminophenyl)-2-phenyloxazole-4-carboxylate (24a)

R_(f)=0.40 (hexane/Et₂O=1:2, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm8.22-8.11 (m, 1H), 8.15 (d, J=7.8 Hz, 1H), 7.54-7.41 (m, 4H), 7.31 (td,J=8.7 and 1.5 Hz, 1H), 6.87 (t, J=7.5 Hz, 1H), 6.84 (d, J=8.1 Hz, 1H),4.42 (q, J=7.2 Hz, 2H), 4.25 (bs, 2H), 1.38 (t, J=7.2 Hz, 3H). LRMS(ESI) m/z 309.3 [M+H]⁺, 331.2 [M+Na]⁺, and 347.0 [M+K]⁺. HRMS (ESI) m/zcalculated for C₁₈H₁₆N₂O₃Na⁺ [M+Na]⁺: 331.1053; found: 331.1051.

(3) Ethyl 5-(2-aminophenyl)-2-(4-chlorophenyl)oxazole-4-carboxylate(24c)

R_(f)=0.23 (hexane/Et₂O=1:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.20(d, J=8.1 Hz, 1H), 8.14 (dd, J=5.4 and 8.9 Hz, 2H), 7.83-7.65 (m, 3H),7.19 (t, J=8.7 Hz, 2H), 4.44 (q, J=7.2 Hz, 2H), 1.29 (t, J=7.2 Hz, 3H);¹³C NMR (75 MHz, CDCl₃) δ ppm 166.3, 161.3, 160.6, 151.0, 148.5, 132.9,132.6, 131.3, 130.2, 129.3, 129.2, 124.9, 122.6, 122.5, 122.5, 116.3,116.0, 61.6, 14.0. LRMS (ESI) m/z 343.1 [M+H]⁺, 365.1 [M+Na]⁺. HRMS(ESI) m/z calculated for C₁₈H₁₅ClN₂O₃Na⁺ [M+Na]⁺: 365.0663; found:365.0678

(4) Ethyl5-(2-aminophenyl)-2-(4-(trifluoromethyl)phenyl)oxazole-4-carboxylate(24d)

R_(f)=0.29 (CH₂Cl₂-MeOH=20:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.25(d, J=8.1 Hz, 2H), 7.74 (d, J=8.1 Hz, 2H), 7.44 (t, J=7.8 Hz, 1H), 7.31(t, J=7.8 Hz, 1H), 6.86 (t, J=7.5 Hz, 1H), 6.83 (d, J=7.8 Hz, 1H), 4.40(q, J=7.2 Hz, 2H), 4.17 (s, 2H), 1.29 (t, J=7.2 Hz, 3H); ¹³C NMR (75MHz, CDCl₃) δ ppm 162.0, 159.2, 155.1, 132.0, 131.6, 127.1, 126.0,125.9, 118.2, 116.9, 112.4, 61.7, 14.2. LRMS (ESI) m/z 356.8 [M+H]⁺ and398.9 [M+Na]⁺. HRMS (ESI) m/z calculated for C₁₈H₁₆F₃N₂O₃ ⁺ [M+Na]⁺:377.1108; found: 377.1094.

(5) Ethyl 5-(2-aminophenyl)-2-(4-methoxyphenyl)oxazole-4-carboxylate(24e)

R_(f)=0.28 (CH₂Cl₂-MeOH=20:1, v/v). ¹H NMR (300 MHz, CDCl₃) δ ppm 8.07(d, J=8.7 Hz, 2H), 7.44 (d, J=7.5 Hz, 1H), 7.28 (t, J=8.1 Hz, 2H), 6.98(d, J=8.7 Hz, 1H), 6.85 (t, J=7.5 Hz, 1H), 6.81 (d, J=7.8 Hz, 1H), 4.39(q, J=7.2 Hz, 2H), 4.18 (bs, 2H), 1.34 (t, J=7.2 Hz, 3H); ¹³C NMR (75MHz, CDCl₃) δ ppm 162.8, 162.3, 157.8, 151.9, 138.4, 130.2, 129.3,122.7, 121.6, 119.1, 115.4, 114.4, 111.6, 61.4, 55.4, 14.2. LRMS (ESI)m/z 339.1 [M+H]⁺ and 361.1 [M+Na]⁺. HRMS (ESI) m/z calculated forC₁₉H₁₉N₂O₄+[M+H]⁺: 339.1339; found: 339.1452.

(e) Synthesis of Quinolinone Derivatives Through Cyclization (1)2-(4-fluorophenyl)oxazolo[4,5-c]quinolin-4(5H)-one (25b)

A mixture of 24b (95 mg, 0.27 mmol), DME (7 mL) and a 2 M potassiumcarbonate solution (0.5 mL, 1.0 mmol) was heated to reflux for 12 h.Thereafter, the solid was collected by filtration, washed with coldEtOH, and dried under vacuum to afford 25b (45 mg, 60%).

R_(f)=0.18 (CH₂Cl₂-MeOH=1:1, v/v). ¹H NMR (300 MHz, DMSO-d₆) δ ppm 11.19(bs, 1H), 7.40 (d, J=9.0 Hz, 1H), 7.38 (d, J=9.0 Hz, 1H), 7.14 (d, J=7.8Hz, 1H), 6.71 (t, J=7.2 Hz, 1H), 6.35 (d, J=7.2 Hz, 1H), 6.59 (d, J=8.7Hz, 1H), 6.47 (t, J=7.8 Hz, 1H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 166.3,161.3, 160.6, 151.0, 148.5, 132.9, 132.6, 131.3, 130.2, 129.3, 129.2,124.9, 122.6, 122.5, 122.5, 116.3, 116.0, 61.6, 14.0. LRMS (ESI) m/z302.7 [M+Na]⁺ and 318.8 [M+K]⁺. HRMS (ESI) m/z calculated forC₁₆H₁₀FN₂O₂ ⁺ [M+H]⁺: 281.0721; found: 281.0713.

(2) 2-Phenyloxazolo[4,5-c]quinolin-4(5H)-one (25a)

R_(f)=0.30 (CH₂Cl₂-MeOH=20:1, v/v). ¹H NMR (300 MHz, DMSO-d₆) δ ppm12.08 (bs, 1H), 8.29-8.18 (m, 2H), 8.05 (d, J=7.8 Hz, 1H), 7.72-7.56 (m,4H), 7.52 (d, J=8.1 Hz, 1H), 7.37 (t, J=7.8 Hz, 1H); ¹³C NMR (75 MHz,CDCl₃) δ ppm 166.3, 161.3, 160.6, 151.0, 148.5, 132.9, 132.6, 131.3,130.2, 129.3, 129.2, 124.9, 122.6, 122.5, 122.5, 116.3, 116.0, 61.6,14.0. LRMS (ESI) m/z 262.8 [M+H]⁺, 285.2 [M+Na]⁺, and 300.6 [M+K]⁺. HRMS(ESI) m/z calculated for C₁₆H₁₁N₂O₂ ⁺[M+H]⁺: 263.0815; found: 263.0845.

(3) 2-(4-Chlorophenyl)oxazolo[4,5-c]quinolin-4(5H)-one (25c)

R_(f)=0.32 (CH₂Cl₂-MeOH=20:1, v/v). ¹H NMR (300 MHz, DMSO-d₆) δ ppm 8.24(d, J=8.4 Hz, 2H), 8.01 (d, J=7.5 Hz, 1H), 7.02 (t, J=8.7 Hz, 2H), 7.56(d, J=7.5 Hz, 1H), 7.51 (t, J=8.4 Hz, 1H), 7.31 (t, J=7.5 Hz, 1H); ¹³CNMR (75 MHz, CDCl₃) δ ppm 166.3, 161.3, 160.6, 151.0, 148.5, 132.9,132.6, 131.3, 130.2, 129.3, 129.2, 124.9, 122.6, 122.5, 122.5, 116.3,116.0, 61.6, 14.0. LRMS (ESI) m/z 335.2 [M+K]⁺. HRMS (ESI) m/zcalculated for C₁₆H₉ClN₂O₂Na⁺[M+Na]⁺: 319.0245; found: 319.0276.

(4) 2-(4-(Trifluoromethyl)phenyl)oxazolo[4,5-c]quinolin-4(5H)-one (25d)

R_(f)=0.38 (CH₂Cl₂-MeOH=20:1, v/v). ¹H NMR (300 MHz, DMSO-d₆) δ ppm 8.44(d, J=8.1 Hz, 2H), 8.08 (d, J=7.8 Hz, 1H), 8.03 (d, J=8.4 Hz, 2H), 7.63(t, J=7.2 Hz, 1H), 7.53 (d, J=8.1 Hz, 1H), 7.38 (t, J=7.5 Hz, 1H); ¹³CNMR (75 MHz, CDCl₃) δ ppm 166.3, 161.3, 160.6, 151.0, 148.5, 132.9,132.6, 131.3, 130.2, 129.3, 129.2, 124.9, 122.6, 122.5, 122.5, 116.3,116.0, 61.6, 14.0. LRMS (ESI) m/z 353.5 [M+Na]⁺ and 369.0 [M+K]⁺. HRMS(ESI) m/z calculated for C₁₇H₁₀F₃N₂O₂ ⁺ [M+H]⁺: 331.0689; found:331.0682.

(5) 2-(4-Methoxyphenyl)oxazolo[4,5-c]quinolin-4(5H)-one (25e)

R_(f)=0.28 (CH₂Cl₂-MeOH=20:1, v/v). ¹H NMR (300 MHz, DMSO-d₆) δ ppm11.18 (bs, 1H), 7.38-7.31 (m, 1H), 7.33 (d, J=9.0 Hz, 1H), 7.19 (dd,J=1.2 and 8.1 Hz, 1H), 6.76 (td, J=8.4 and 1.5 Hz, 1H), 6.68 (d, J=9.0Hz, 1H), 6.53 (td, J=8.1 and 1.2 Hz, 1H), 6.41-6.33 (m, 1H), 6.35 (d,J=9.0 Hz, 1H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 166.3, 161.3, 160.6, 151.0,148.5, 132.9, 132.6, 131.3, 130.2, 129.3, 129.2, 124.9, 122.6, 122.5,122.5, 116.3, 116.0, 61.6, 14.0. LRMS (ESI) m/z 293.1 [M+H]⁺, 314.9[M+Na]⁺, and 331.0 [M+K]⁺. HRMS (ESI) m/z calculated forC₁₇H₁₃N₂O₃+[M+H]⁺: 293.0921; found: 293.0919.

(f) Synthesis of Quinolinone Derivatives as Final Compounds ThroughAlkylation (1)5-(2-(dimethylamino)ethyl)-2-(4-fluorophenyl)oxazolo[4,5-c]quinolin-4(5H)-one(3)

DMF (5 mL) was added to 25b (43 mg, 0.153 mmol),2-bromo-N,N-dimethylethanaminehydrobromide (80 mg, 0.31 mmol, 1.2 eq.),and potassium carbonate (63 mg, 0.46 mmol, 3.0 eq.). The mixture wasstirred at 130° C. for 3 h. After cooling to room temperature, thereaction mixture was evaporated under vacuum to remove the solvent andpurified by silica gel column chromatography using Et₂O-MeOH (10:1 to3:1, v/v) as the eluent, giving the compound of Formula 3.

Yield: 39%. R_(f)=0.15 (EtO₂-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.33-8.22 (m, 1H), 8.29 (dd, J=5.4 and 8.7 Hz, 1H), 8.03 (dd, J=1.5and 8.0 Hz, 1H), 7.64 (td, J=7.2 and 1.2 Hz, 1H), 7.57 (d, J=8.4 Hz,1H), 7.38 (t, J=6.9 Hz, 1H), 7.22 (t, J=8.4 Hz, 1H), 7.27-7.16 (m, 1H),4.58 (t, J=7.8 Hz, 2H), 2.66 (t, J=7.8 Hz, 2H), 2.41 (s, 6H); ¹³C NMR(75 MHz, CDCl₃) δ ppm 166.5, 163.1, 161.7, 157.3, 152.3, 130.5, 129.8,129.7, 129.6, 122.8, 122.8, 121.9, 116.4, 116.1, 115.4, 111.6, 56.3,45.9, 40.8, 29.7, 22.7. LRMS (ESI) m/z 352.2 [M+H]⁺ and 374.0 [M+Na]⁺.HRMS (ESI) m/z calculated for C₁₈H₁₉FN₃O₂+[M+H]⁺: 352.1456; found:352.1496.

(2) 5-(2-(dimethylamino)ethyl)-2-phenyloxazolo[4,5-c]quinolin-4(5H)-one(2)

Yield: 39%. R_(f)=0.25 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.34-8.27 (m, 2H), 8.07 (dd, J=1.5 and 7.8 Hz, 1H), 7.69-7.51 (m,5H), 7.38 (td, J=7.2 and 1.2 Hz, 1H), 4.59 (t, J=7.5 Hz, 2H), 2.68 (t,J=7.5 Hz, 2H), 2.41 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 162.6, 157.4,152.3, 137.9, 131.6, 130.5, 130.0, 129.0, 127.4, 126.5, 122.6, 122.0,115.4, 111.8, 56.3, 45.9, 40.9. LRMS (ESI) m/z 333.4 [M+H]⁺ and 355.3[M+Na]⁺. HRMS (ESI) m/z calculated for C₂₀H₂₀N₃O₂ ⁺ [M+H]⁺: 334.1550;found: 334.1557.

(3)2-(4-chlorophenyl)-5-(2-(dimethylamino)ethyl)oxazolo[4,5-c]quinolin-4(5H)-one(4)

Yield: 26%. R_(f)=0.17 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.24 (d, J=8.7 Hz, 2H), 8.04 (d, J=8.1 Hz, 1H), 7.96 (d, J=8.4 Hz,1H), 7.68 (td, J=7.5 and 1.5 Hz, 1H), 7.53 (d, J=8.7 Hz, 2H), 7.42 (t,J=7.8 Hz, 1H), 4.43 (t, J=5.1 Hz, 2H), 2.86 (t, J=5.1 Hz, 2H), 2.41 (s,6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 151.6, 138.1, 136.9, 130.6, 129.4,129.0, 128.7, 127.5, 123.5, 121.44, 113.7, 109.5, 57.3, 53.4, 45.8,29.7. LRMS (ESI) m/z 368.0 [M+H]⁺ and 390.2 [M+Na]⁺. HRMS (ESI) m/zcalculated for C₂₀H₁₉ClN₃O₂ ⁺ [M+H]⁺: 368.1160; found: 368.1202.

(4)5-(2-(dimethylamino)ethyl)-2-(4-(trifluoromethyl)phenyl)oxazolo[4,5-c]quinolin-4(5H)-one(5)

Yield: 37%. R_(f)=0.13 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.43 (d, J=8.1 Hz, 2H), 8.09 (dd, J=1.2 and 7.8 Hz, 1H), 7.81 (d,J=8.4 Hz, 2H), 7.68 (dddd, J=1.5, 7.2, and 8.7 Hz, 1H), 7.60 (d, J=8.4Hz, 1H), 7.41 (td, J=7.8 and 0.9 Hz, 1H), 4.60 (t, J=7.5 Hz, 2H), 2.68(t, J=7.8 Hz, 2H), 2.42 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 161.0,157.3, 152.8, 138.1, 131.0, 127.7, 126.1, 126.1, 126.0, 126.0, 122.8,122.1, 115.5, 111.5, 56.3, 45.9, 37.6. LRMS (ESI) m/z 402.1 [M+H]⁺ and424.1 [M+Na]⁺. HRMS (ESI) m/z calculated for C₂₁H₁₉F₃N₃O₂+[M+H]⁺:402.1424; found: 402.1426.

(5)5-(2-(dimethylamino)ethyl)-2-(4-methoxyphenyl)oxazolo[4,5-c]quinolin-4(5H)-one(6)

Yield: 18%. R_(f)=0.08 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.24 (d, J=9.0 Hz, 2H), 8.04 (d, J=8.4 Hz, 1H), 7.64-7.60 (m, 2H),7.40-7.33 (m, 1H), 7.04 (d, J=8.7 Hz, 2H), 4.62 (t, J=7.5 Hz, 2H), 3.09(s, 3H), 2.72 (t, J=7.8 Hz, 2H), 2.44 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δppm 162.8, 162.4, 157.5, 152.0, 137.6, 130.1, 129.9, 129.4, 122.6,121.8, 119.1, 115.4, 114.4, 111.8, 56.0, 55.5, 45.6, 40.5, 29.7. LRMS(ESI) m/z 364.1 [M+H]⁺ and 387.1 [M+Na]⁺. HRMS (ESI) m/z calculated forC₂₁H₂₂N₃O₃ ⁺ [M+H]⁺: 364.1656; found: 364.1694.

(6) N,N-dimethyl-2-((2-phenyloxazolo[4,5-c]quinolin-4-yl)oxy)ethanamine(7)

Yield: 9%. R_(f)=0.36 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.38-8.31 (m, 2H), 8.16 (dd, J=0.9 and 8.1 Hz, 1H), 7.96 (d, J=8.4Hz, 1H), 7.66 (td, J=7.2 and 1.5 Hz, 1H), 7.59-7.48 (m, 4H), 4.84 (t,J=6.0 Hz, 2H), 2.94 (t, J=6.0 Hz, 1H), 2.42 (s, 6H); ¹³C NMR (75 MHz,CDCl₃) δ ppm 163.0, 154.7, 153.9, 144.2, 131.6, 129.1, 129.0, 127.9,127.6, 126.7, 126.5, 124.7, 120.3, 114.7, 64.1, 57.9, 45.9. LRMS (ESI)m/z 333.9 [M+H]⁺ and 356.9 [M+Na]⁺. HRMS (ESI) m/z calculated forC₂₀H₂₀N₃O₂+[M+H]⁺: 334.1550; found: 334.1552.

(7)2-((2-(4-fluorophenyl)oxazolo[4,5-c]quinolin-4-yl)oxy)-N,N-dimethylethanamine(8)

Yield: 9%. R_(f)=0.30 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.41-8.28 (m, 11H), 8.29 (dd, J=5.4 and 9.0 Hz, 1H), 8.14 (d, J=8.1Hz, 1H), 7.96 (d, J=8.1 Hz, 1H), 7.67 (ddd, J=1.5, 7.1, and 8.4 Hz, 1H),7.52 (td, J=8.0 and 0.9 Hz, 1H), 7.29-7.18 (m, 2H), 4.83 (t, J=6.0 Hz,2H), 2.93 (t, J=6.0 Hz, 2H), 2.41 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm166.6, 163.2, 161.8, 154.6, 153.9, 144.2, 129.9, 129.8, 128.0, 126.4,124.8, 123.0, 123.0, 120.2, 116.4, 116.1, 114.6, 64.1, 57.9, 45.8, 29.7.

LRMS (ESI) m/z 352.2 [M+H]⁺ and 374.0 [M+Na]⁺. HRMS (ESI) m/z calculatedfor C₂₀H₁₉FN₃O₂ ⁺ [M+H]⁺: 352.1456; found: 352.1498.

(8)2-((2-(4-chlorophenyl)oxazolo[4,5-c]quinolin-4-yl)oxy)-N,N-dimethylethanamine(9)

Yield: 8%. R_(f)=0.28 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.27 (d, J=8.4 Hz, 2H), 8.14 (d, J=8.1 Hz, 1H), 7.96 (d, J=8.1 Hz,1H), 7.67 (t, J=7.5 Hz, 1H), 7.53 (d, J=8.1 Hz, 3H), 4.84 (t, J=5.1 Hz,2H), 2.94 (t, J=5.1 Hz, 2H), 2.42 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm161.7, 154.6, 153.9, 144.3, 138.0, 129.4, 129.3, 128.4, 128.0, 126.4,125.2, 124.8, 120.2, 114.6, 64.8, 57.9, 53.4, 45.8, 31.9. LRMS (ESI) m/z368.2 [M+H]⁺ and 390.2 [M+Na]⁺. HRMS (ESI) m/z calculated forC₂₀H₁₉ClN₃O₂ ⁺ [M+H]⁺: 368.1160; found: 368.1195.

(9)N,N-dimethyl-2-((2-(4-(trifluoromethyl)phenyl)oxazolo[4,5-c]quinolin-4-yl)oxy)ethanamine(10)

Yield: 10%. R_(f)=0.38 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.47 (d, J=8.1 Hz, 2H), 8.18 (d, J=7.8 Hz, 1H), 7.98 (d, J=8.4 Hz,1H), 7.82 (d, J=8.4 Hz, 2H), 7.69 (td, J=7.2 and 1.5 Hz, 1H), 7.54 (td,J=7.2 and 1.2 Hz, 1H), 4.49 (t, J=6.3 Hz, 2H), 2.94 (t, J=6.0 Hz, 2H),2.42 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 166.1, 154.7, 154.1, 144.4,129.8, 129.5, 128.0, 127.8, 126.4, 126.0, 126.0, 124.9, 120.4, 114.5,64.2, 59.6, 45.9. LRMS (ESI) m/z 402.1 [M+H]⁺ and 424.1 [M+Na]⁺. HRMS(ESI) m/z calculated for C₂₁H₁₉F₃N₃O₂+[M+H]⁺: 402.1424; found: 402.1437.

(10)2-((2-(4-methoxyphenyl)oxazolo[4,5-c]quinolin-4-yl)oxy)-N,N-dimethylethanamine(11)

Yield: 15%. R_(f)=0.18 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.26 (d, J=8.1 Hz, 2H), 8.16 (d, J=6.3 Hz, 1H), 7.96 (d, J=8.4 Hz,1H), 7.66 (dd, J=6.9 and 14.3 Hz, 1H), 7.54 (dd, J=7.8 and 16.2 Hz, 1H),7.05 (d, J=8.7 Hz, 2H), 4.86 (t, J=6.3 Hz, 2H), 3.91 (s, 3H), 3.01 (t,J=6.3 Hz, 2H), 2.47 (s, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 163.0, 162.5,154.6, 153.6, 144.0, 129.4, 128.8, 127.9, 126.5, 124.7, 120.2, 119.2,114.8, 114.5, 63.7, 57.8, 55.5, 45.6, 30.4. LRMS (ESI) m/z 364.5 [M+H]⁺and 387.5 [M+Na]⁺. HRMS (ESI) m/z calculated for C₂₁H₂₂N₃O₃ ⁺ [M+H]⁺:364.1656; found: 364.1693.

(11)5-(2-(diethylamino)ethyl)-2-(4-fluorophenyl)oxazolo[4,5-c]quinolin-4(5H)-one(12)

Yield: 14%. R_(f)=0.39 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.35-8.26 (m, 1H), 8.30 (dd, J=5.4 and 8.7 Hz, 1H), 8.03 (d, J=7.5Hz, 1H), 7.66-7.59 (m, 2H), 7.43-7.34 (m, 1H), 7.22 (t, J=8.7 Hz, 2H),4.55 (t, J=7.8 Hz, 2H), 2.79 (t, J=7.8 Hz, 2H), 2.68 (dd, J=7.2 and 14.4Hz, 4H), 1.09 (t, J=7.2 Hz, 6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 166.5,163.2, 161.7, 157.4, 152.3, 138.0, 130.5, 129.8, 129.7, 129.6, 122.8,122.8, 122.6, 121.8, 116.4, 116.1, 115.6, 111.6, 50.0, 47.6, 41.2, 12.0.LRMS (ESI) m/z 380.3 [M+H]⁺. HRMS (ESI) m/z calculated for C22H₂₃FN₃O₂⁺[M+H]⁺: 380.1769; found: 380.1817.

(12)N,N-diethyl-2-((2-(4-fluorophenyl)oxazolo[4,5-c]quinolin-4-yl)oxy)ethanamine(13)

Yield: 8%. R_(f)=0.51 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.38-8.31 (m, 1H), 8.35 (dd, J=5.4 and 9.0 Hz, 1H), 8.14 (d, J=7.8Hz, 1H), 7.96 (d, J=8.4 Hz, 1H), 7.66 (ddd, J=1.5, 7.1, and 8.4 Hz, 1H),7.52 (t, J=7.5 Hz, 1H), 7.28-7.19 (m, 2H), 4.81 (t, J=6.9 Hz, 2H), 3.06(t, J=6.9 Hz, 2H), 2.72 (dd, J=7.2 and 14.4 Hz, 4H), 1.14 (t, J=7.2 Hz,6H); ¹³C NMR (75 MHz, CDCl₃) δ ppm 166.6, 163.2, 161.8, 154.6, 153.8,144.2, 129.9, 129.8, 129.1, 128.0, 126.4, 124.7, 123.0, 123.0, 120.2,116.5, 116.2, 114.6, 64.2, 51.0, 47.7, 29.7, 11.8. LRMS (ESI) m/z 380.3[M+H]⁺. HRMS (ESI) m/z calculated for C₂₂H₂₃FN₃O₂ ⁺ [M+H]⁺: 380.1769;found: 380.1814.

(13)5-(2-(diethylamino)ethyl)-2-(4-(trifluoromethyl)phenyl)oxazolo[4,5-c]quinolin-4(5H)-one(14)

Yield: 27%. R_(f)=0.39 (Et₂O-MeOH=5:1, v/v). ¹H NMR (300 MHz, CDCl₃) δppm 8.44 (d, J=8.1 Hz, 2H), 8.08 (d, J=7.8 Hz, 1H), 7.81 (d, J=8.1 Hz,2H), 7.71-7.60 (m, 2H), 7.70 (t, J=7.8 Hz, 1H), 4.56 (t, J=7.8 Hz, 2H),2.80 (t, J=8.1 Hz, 2H), 2.69 (dd, J=7.2 and 14.4 Hz, 4H), 1.09 (t, J=7.2Hz, 6H).

(14)N,N-diethyl-2-((2-(4-(trifluoromethyl)phenyl)oxazolo[4,5-c]quinolin-4-yl)oxy)ethanamine(15)

Yield: 50%. R_(f)=0.51 (Et₂O-MeOH=5:1, v/v). 1HNMR (300 MHz, CDCl₃) δppm 8.45 (d, J=8.1 Hz, 2H), 8.15 (d, J=8.7 Hz, 1H), 7.95 (d, J=8.4 Hz,1H), 7.80 (d, J=8.4 Hz, 2H), 7.68 (ddd, J=1.5, 7.1, and 8.4 Hz, 1H),7.53 (t, J=7.2 Hz, 1H), 4.81 (t, J=6.9 Hz, 2H), 3.06 (t, J=7.2 Hz, 2H),2.72 (dd, J=7.2 and 14.4 Hz, 4H), 1.09 (t, J=7.2 Hz, 6H).

Experimental Example 1: Evaluation of Inhibitory Efficacy on TSLP-TSLPRInteraction by ELISA

In this example, an investigation was made as to whether the inventivequinolinone derivative compounds directly inhibit the binding of TSLP toTSLP receptor. To this end, home-made FLAG-TSLP protein and TSLPR-Hisprotein were used to establish an ELISA-based TSLP-TSLPR interactionassay for measuring reactions with HRP. The inhibitory activities of theinventive compounds were evaluated by the interaction assay.

ELISA assay was conducted using an Ni-NTA HisSorb plate (Qiagen,Germany). 100 μL of a solution of TSLPR C-terminally tagged withhexahistidine (His) was dispensed into each well of the plate andincubated at room temperature for 2 h. After each well was washed twicewith 200 μL of PBS and 0.05% Tween-20, 100 μL of a solution of each ofthe inventive compounds as a sample and 100 μL of TSLP N-terminallytagged with FLAG was dispensed into the well. After incubation at 4° C.overnight, each well was washed twice and treated with 100 μL of ablocking buffer (PBS-0.05% Tween 20 and 1% nonfat dry milk). The platewas washed twice, coated with 100 μL of a monoclonal anti-FLAGhorseradish peroxidase antibody (Sigma-Aldrich Co., USA) for 2 h, andincubated at room temperature. Then, each well was washed 5 times,treated with 200 μL of a solution of o-phenylenediaminedihydrochloride(Sigma-Aldrich Co., USA), incubated for 30 min, and added with 1 N HCl.

Optical densities (ODs) were measured at 450 nm using a microplatespectrophotometer. The inhibitory effect of the compound on TSLP-TSLPRinteraction was calculated using the following equation:Inhibition (inhibitory effect) (%)=(1−OD_(sample)/OD_(control))×100

The results are shown in Table 1.

TABLE 1 Inhibition (%) Compound 0.1 mM 0.3 mM 1 mM 2 26.7 49.0 66.9 317.7 27.5 N/D 4 28.8 46.6 N/D 5 23.2 29.6 N/D 6 25.8 54.7 N/D 7 27.854.6 57.2 FLAG tag-free 40.7 TSLP (1 mg/mL) N/D: Not determined

Experimental Example 2: Evaluation of Inhibitory Efficacy on IL-6Secretion by IL-33 Inhibition

First, HMC-1, a human mast cell line, was cultured until an appropriatenumber of cells was maintained in a 96-well plate. Mixtures of IL-33 anddifferent concentrations (0.3, 3, and 30 μM) of each of the inventivecompounds were treated on cells, incubated at 37° C. for 24 h, andcentrifuged. The supernatants were analyzed for IL-6 secretion using anIL-6 ELISA kit (Biolegend, USA). The results are shown in Table 2.

TABLE 2 Inhibition of IL-6 secretion (%) Compound 0.03 μM 0.3 μM 3 μM 2— −11.8 3 — 29.8 4 22.5 29.7 5 30.4 28.2 6 — −57.8 7 — 35.6 10 32.7 49.351.3 11 — −216.4 IL-33 only 0 Negative (−) values indicate increasedIL-6 secretion

Taken together, these results demonstrate that the inventive quinolinonederivatives effectively inhibit TSLP and IL-33, key cytokines inducingallergic diseases such as asthma or atopic dermatitis, from binding totheir receptors. In conclusion, the inventive quinolinone derivativescan be used to fundamentally prevent and treat allergic diseases such asasthma or atopic dermatitis due to their ability to inhibit TSLP- orIL-33-mediated signal transduction.

Experimental Example 3: House Dust Mite (HDM)-Induced Allergic AirwayInflammation Mouse Models

Splenocytes were isolated from DO11.10 mice and ˜1×10⁶CD4T cells weretransferred to pure BALB/c mice by intravenous injection. For 3 daysfrom the next day, 100 μg of a mixture of HDM (Dermatophagoidesfarinae50 μg, Dermatophagoidespteronyssinus 50 μg, Greer Laboratories, Inc.,USA) and OVA (Sigma-Aldrich Co., USA) was administered intranasally tothe mice. From day 2 after administration, the mice were treated with200 μg of each of the inventive compounds or PBS (for control mice)three times daily through the intraperitoneal route. After the mice wereeuthanized on day 11, the numbers of total cells and eosinophils in theairways were checked. The treatment schedule with the Formula 5 (KB1517) and Formula 10 (KB 1518) (200 μg each) in house dust mite(HDM)-induced allergic airway inflammation mouse models is shown in FIG.8. The results are shown in FIGS. 4 and 5.

As can be seen from FIGS. 4 and 5, the numbers of total cells andeosinophils in the airways of the mice treated with the inventivecompounds of Formula 5 (KB-1517) and Formula 10 (KB-1518) (200 μg each)were significantly reduced than those in the untreated controls. Theseresults lead to the conclusion that the inventive quinolinonederivatives can be used to fundamentally prevent and treat allergicdiseases such as asthma atopic dermatitis.

Experimental Example 4: 2D NMR Spectrometry

Measurements were done using a Bruker 600 MHz NMR spectrometer(triple-resonance, pulsed field gradient probe (Bruker, Germany)) at 25°C. 2D ¹H-¹⁵NHSQC spectra of IL-33 were measured in the same molar ratiowith/without each of the compounds. The 2D data were processed andanalyzed using the TopSpin 3.1 program (Bruker, Germany).

Chemical shift perturbations (CSPs) were calculated using the followingequation:

${CSP} = \sqrt{\left( {\Delta\delta}_{1\; H} \right)^{2} + \frac{\left( {\Delta\delta}_{15\; N} \right)^{2}}{5}}$

where Δδ_(1H) and Δδ_(15N) indicate H and ¹⁵N chemical shiftdifferences, respectively.

Experimental Example 5: IL-33 Protein Expression and Purification

IL-33 was cloned into an expression vector pPROEX, an N-terminal His-tagfusion protein, in E. coli BL21 (DE3). When the cell density (OD₆₀₀)reached 0.6, cells were induced with 0.5 mMisopropyl-1-D-thiogalactoside (IPTG) and were allowed to further grow at20° C. overnight. To obtain uniformly labeled ⁵N IL-33, the bacterialcells were grown in M9 minimal medium containing ¹⁵N NH₄Cl. Cells wereharvested and resuspended in lysis buffer (0.1 M Tris pH 7.4, 0.3 MNaCl, 1 mM β-mercaptoethanol, 0.1% TritonX100, and 0.1 mMphenylmethylsulfonyl fluoride). Cells were lysed by sonication in an icebath. The cell lysate was centrifuged at 10000×g and 4° C. for 25 min.The pellets were discarded and the supernatant was eluted with 50 mMsodium phosphate (pH 7.4), 300 mM sodium chloride, and 1 mMβ-mercaptoethanol from a 5 mL HisPur cobalt resin column (ThermoScientific Inc.). Thereafter, the buffer was exchanged with 50 mM sodiumphosphate (pH 7.4), 300 mM sodium chloride, 1 mM β-mercaptoethanol, and250 mM imidazole. The fusion protein was cleaved with TEV protease indialysis buffer (Tris 20 mM (pH 7.4), sodium chloride 300 mM) at 4° C.The IL-33-containing mixture was loaded onto a HisPur cobalt resincolumn and washed with 0.1 M Tris (pH 7.4), 0.3 M sodium chloride, and 1mM β-mercaptoethanol. Thereafter, the bound protein was eluted with 0.1M Tris (pH 7.4), 0.3 M sodium chloride and 1 mM p-mercaptoethanol (20mL). The eluate was loaded onto a Superdex S75 gel filtration column(16/60 GE Healthcare) equilibrated with a buffer containing 20 mM sodiumphosphate (pH 6.8), 100 mM sodium chloride, and 5 mM BME to obtain pureIL-33 protein.

INDUSTRIAL APPLICABILITY

The quinolinone derivative compounds of the present invention caneffectively suppress inflammatory responses of an allergic disease suchas asthma or atopic dermatitis. In addition, the pharmaceuticalcomposition of the present invention can be used to fundamentallyprevent or treat various allergic diseases such as asthma or atopicdermatitis.

The invention claimed is:
 1. A quinolinone derivative represented byFormula 1a or Formula 1 b:

wherein R₁ and R2 are identical to or different from each other and areeach independently selected from C₁-C₄ alkyl groups, and —Z-alkyl(wherein Z is selected from O, S, or is —(CH₂)_(m)— wherein m is aninteger from 0 to 5), X and Y are each independently selected fromhydrogen, halogen groups, a hydroxyl group, an amino group, a nitro Agroup, a cyano group, a trifluoromethyl group, CO—R′ (wherein R′ is aC₁-C₄ alkyl group), NHR′ (wherein R′ is a C₁-C₄ alkyl group), NR′R′(wherein each R′ is independently a C₁-C₄alkyl group), NHCOR′ (whereinR′ is a C₁-C₄ alkyl group), and COOR′ (wherein R′ is a C₁-C₄ alkylgroup), p is an integer from 0 to 4, provided that when p is an integerof 2 or more, the Y groups are identical to or different from eachother, and q is an integer from 0 to 5, provided that when q is aninteger of 2 or more, the X groups are identical to or different fromeach other,

wherein R₁, R₂, X, Y, p, and q are as defined in Formula 1a.
 2. Aquinolinone derivative represented by Formula 1a or Formula 1b selectedfrom the group consisting of the following compounds:


3. A pharmaceutical composition for preventing or treating an allergicdisease comprising the quinolinone derivative of claim 1 as an activeingredient and a pharmaceutically acceptable carrier.